Development of capillary electrophoresis method to measure albumin thiol oxidation in dystrophic humans and animal models of Duchenne muscular dystrophy.

Inflammatory responses evident in many diseases involve the generation of oxidants which can cause oxidant-induced post-translational modifications to proteins. Albumin, the most abundant plasma protein, contains a free thiol group which is susceptible to oxidative modification. We propose that albumin thiol oxidation (AlbOx) could be a useful biomarker to monitor changes in inflammatory activity and oxidative stress. To measure AlbOx in humans and animal models, we developed a fast, sensitive, simple, and reproducible capillary electrophoresis method (CE-AlbOx). This method can analyse total, reversible, and irreversible oxidation of albumin. The method only requires a small volume of sample (<10 μL blood), has an intra/interday variation of <2 %, and has a total run time of 17 min. We validated the usefulness of AlbOx as a biomarker of chronic inflammation by analysing samples from patients with, and animal models of, Duchenne muscular dystrophy (DMD), a disease associated with chronic inflammation. The main findings in this study are (1) dystrophic humans and animals have higher oxidised albumin compared to healthy controls, (2) mouse albumin has two reactive cysteine groups, and (3) our method is the first to quantify the different oxidation states of mouse albumin. In conclusion, we have developed a new method to measure albumin oxidation in humans and animals using capillary electrophoresis. The simple methodology of the CE-AlbOx method makes it advantageous to current methods and can be readily used as a biomarker of inflammation and oxidative stress in both humans and animal models.